# Mechanisms mediating age-dependent inhibition of cerebrovascular MLCK activity and contractility by chronic hypoxia

> **NIH NIH P01** · LOMA LINDA UNIVERSITY · 2020 · $122,763

## Abstract

Cardiovascular instability is a common feature of NICU infants that often leads to compromised cerebral
autoregulation, hypoxic-ischemic brain injury, and intraventricular hemorrhage. Our recent work suggests that
postnatal cardiovascular instability involves depressed function of Myosin Light Chain Kinase (MLCK), the rate-
limiting enzyme responsible for initiation and regulation of vascular contraction. Because rates of mRNA
transcription for MLCK vary little with age and hypoxia, our results implicate changes in mRNA translation,
MLCK degradation, and MLCK activity as the main mechanisms that govern neonatal MLCK function. First,
we will examine effects of micro-RNAs on MLCK translation. Numerous micro-RNAs are induced by hypoxia
and influence contractile protein expression directly through binding to transcripts, and indirectly by influencing
smooth muscle differentiation. To explore these mechanisms we have developed surgical methods that
enable the in vivo adenoviral transfection of pre-term fetal lambs, in utero. This approach offers
unprecedented opportunities to explore the molecular roles of micro-RNAs in fetal responses to hypoxic stress,
particularly as related to regulation of MLCK function. Second, we will examine the roles of ubiquitination and
protein degradation in fetal vascular responses to hypoxia. Despite the recognized importance of
ubiquitination, it has not been studied in fetal lambs, their cerebral arteries or their responses to hypoxia. Our
findings demonstrate that expression of some ubiquitin ligases is age-dependent and for others is potently
upregulated by chronic hypoxia. These results advance the novel idea that changes in protein degradation are
intimately involved in fetal vascular adaptation to chronic hypoxia. Third, we will examine effects of hypoxia on
MLCK activity, in situ. Using novel methods to measure high-speed transients in cytosolic calcium and myosin
light chain phosphorylation in whole arteries, we have found that MLCK velocity is enhanced by chronic
hypoxia in fetal but not adult arteries. Our confocal methods further suggest that colocalization of MLCK with
its substrate is stronger in fetal than adult arteries, and is significantly altered by chronic hypoxia, suggesting a
new role for MLCK compartmentalization in regulation of fetal cerebrovascular contractility. Overall, further
study of the mechanisms identified by our recent work promises to reveal multiple important new features of
the molecular, cellular, and tissue level regulation of MLCK function, and offers new understanding of how
these mechanisms might be leveraged to improve clinical management of postnatal cardiovascular instability.

## Key facts

- **NIH application ID:** 9855057
- **Project number:** 5P01HD083132-05
- **Recipient organization:** LOMA LINDA UNIVERSITY
- **Principal Investigator:** William J. Pearce
- **Activity code:** P01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $122,763
- **Award type:** 5
- **Project period:** — → 2023-02-28

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9855057

## Citation

> US National Institutes of Health, RePORTER application 9855057, Mechanisms mediating age-dependent inhibition of cerebrovascular MLCK activity and contractility by chronic hypoxia (5P01HD083132-05). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/9855057. Licensed CC0.

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